Chromatography Forum

Hi All,

I''ve been given a task of analysing lecithin samples by HPLC.

I was given this paper as a starting point: http://www.springerlink.com/content/x68h782u17148p38/

Essentially it is normal phase with a quite old (and obscure column - porasil) - I'm aiming to just subsitute this column with a modern equivalent - Luna bare Silica (2) - will this perform ok do people think? Will I still need water in the mobile phase as per the method in the journal?

I'm not concerened about separating out the individual lecithin components by rather just assessing for "total lecithin" content in some API samples. Not sure what levles will be present, but lets say as an estimate I need to get to be able to see 1% w/w - so need a relatively low LoQ.

I'm also goint to try and keep it simple with respect to dteectors - i.e. UV down at 210 nm - so we can easily transfer it for routine analysis by chemists.

Jeff Hurst is a very smart practicing chromatographer.

There is nothing wrong with using water as a mobile phase component in normal phase. It gets around all the lengthy equilibration problems that one has with low water content. You can use a more modern silica, but I would start off with the mobile phases that he was working with and adjust as needed for the different silica.

I would say this is a HILIC application not normal phase, but I agree with Uwe try the same conditions using a silica HILIC column. And if that does not work you should naturally use our zwitterionic HILIC column.

I have used a Waters Symmetry C8 column (3.0 x 150 mm, 5um) with good results separating the components of Soy lecithin. Starting conditions are 82% MeOH with 0.1% TFA, 1.5 ml/min, 55 C, and eluted with a non-linear gradient (no 8 on Waters equipment) in 15 min. UV detection at 205 nm.
I have not determined the LoQ, but 10 ul of a 0.1% solution (1 mg/ml) is the lower end of my standard curve. If you use ELS detection you will have higher sensitivity but UV detection was fine for my purposes.
BTW, cholesterol is also separated from Soy lecithin in this set-up.